Central to the pathogenicity of all Salmonellae is their ability to gain access to host cells that are normally non-phagocytic. Despite recent important advances in the understanding of the interaction of Salmonella with host cells, the mechanisms of entry as well as the bacterial determinants that trigger this process remain poorly characterized. We have been studying a Salmonella genetic locus, inv, that is required for bacterial entry into non-phagocytic cells. The extension of the molecular and functional characterization of this locus is the focus of this Grant proposal. It is now clear that the inv locus encodes a dedicated, sec- independent, type-Ill protein secretion system presumably required for the surface presentation of factors necessary for the entry process or for the biogenesis of a supramolecular structure involved in the delivery of entry determinants. This secretion system, which is also present in other bacterial pathogens, is different from the type I sec-independent system exemplified by the export of the E. coli hemolysin or the type II sec-dependent export pathway exemplified by the secretion of pullulanase, an enzyme of the gram negative organism Klebsiella oxytoca. Although proteins whose export depends on the function of type III systems have been identified in several microorganisms (e. g. Shigella Epee's and Yersinia Yops), only one such proteins, InvJ, has been so far identified in Salmonella. Identification of targets of this secretion apparatus is of great interest since it may lead to the identification of putative Salmonella effector molecules that trigger bacterial uptake. Our long term objective is to understand the molecular bases of Salmonella entry. In particular, our specific aims are: 1) to conduct a functional analysis of the protein secretion system encoded in the inv locus; 2) to identify additional targets of this export system; and 3) to investigate their putative effector functions on the host cell.